CN106795490A - The medical composition of nerve degenerative diseases is treated with mitochondria specialized cell - Google Patents
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Abstract
The present invention provides a kind of mitochondria specialized cell processed through Chinese angelica root extract.Present invention also offers medical composition, the medical composition includes mitochondria specialized cell.The medical composition effectively achieves the purpose for treating nerve degenerative diseases.
Description
Cross-Reference to Related Applications
The application of this non-transitory requires No. 62/049,030 that September in 2014 is submitted on the 11st according to 35U.S.C. § 119 (a)
The priority of U.S. Patent application, it passes through reference and is all incorporated into the application.
Technical field
The present invention is related to a kind of medical composition, and particularly with regard to can effectively improve and treat nervus retrogression
Disease, and increase the medical composition that activating cell is divided into the ratio of class neuronal cell.
Background technology
Due to the progress of economic development and health care, the average life span of the mankind rises year by year, population structure it is aging
It is worldwide trend.Shown according to the United Nations's statistics, up to 70.8 hundred million, over-65s population accounts for the whole world to population in the world in 2012
7.9%, it has been aging society (Aging society) defined in the World Health Organization (WHO).Along with aging populations
Increase, the number of nerve degenerative diseases also same rapid growth, the current whole world at least 4,000,000 people suffer from this disease
Disease.However, nerve degenerative diseases are not apt to occur in the elderly, the patient of about half at 60 years old with sequela, second half then in
Fall ill before within 60 years old.Nerve degenerative diseases are a kind of morbid states of the cellular neural unit cell degradation of brain and spinal cord, by
The function of its synapse or its myelin is lost to be caused, and will cause dysfunction, handicapped or death.
Parkinson's disease is a kind of neurogenic disease for more often occurring in the elderly, and age of onset is with 50 to 79
Year is most common.And its pathological characters is the dopaminergic cell degeneration and death in the Nigral Tissue of midbrain, normal Nigral Tissue is interior about
Containing 200,000 dopaminergic neuronal cells, dopamine neuronal can Dopamine Secreted, the specially coordination of department's control motion.These nerves are few
Perhaps degeneration can't cause the uncoordinated of any motion, but when degenerating more than 50%, just start light symptoms occur, including
Limbs tremble, stiff and slow in one's movements.The more, then above-mentioned symptom can be more serious for nerve degeneration, finally to needing what people looked after completely
Invalidity state；Finally may because respiratory tract, the urinary tract, the infection of bedsore and it is dead.
At present, the early stage in Parkinson's disease supplies dopamine neuronal dopamine medicine by drug therapy, so as to
More dopamines are produced to make up the manufacture that the nerve of degeneration is reduced.The Parkinson's disease patient of early stage is for DOPA
The curative effect reaction of medicine is all pretty good.But due to Parkinson's disease for progressive disease, dopamine neuronal always in death,
So that DOPA medicine more can be taken the more, then drug effect is more and more poor, and symptom is just more and more serious.Many patients are in drug administration
After several years, side effects of pharmaceutical drugs, including illusion, nausea, digestive discomfort, or even the involuntary limbs dancing of whole body will occur
Deng.Because drug therapy is in the symptom of later stage uncontrollable severe patient, therefore need to surgically improve the life of patient
Bioplasm amount.Wherein, operative treatment can then be subdivided into following three kinds of forms：(1) otomy is burnt, that is, some cells of brain
Domain is heated to 80 DEG C, and the time is 80 seconds, local nerve cell is lost function, these regions include globus pallidus, Thalamus and
Core etc. under Thalamus；(2) embedded electrode, similar with execution is burnt, its principle is local nerve cell after electrode is powered
Function will be lost；And the operation of (3) stem-cell therapy, its principle is exactly the dopaminergic cell that supplement is degenerated, and will be increased always
It is long, increase intracerebral dopaminergic cell, improve the symptom of Parkinson's disease.
Although the past has document and discloses and can treat Parkinson's disease using stem cell is transplanted, although can improve handkerchief
The symptom of golden Sen Shi diseases, but effect does not reach the degree of healing.After main cause is administered in vivo including stem cell
The ratio that dopamine neuron is divided into after survival rate is not good because of immune response, and stem cell is administered in vivo is relatively low
(Cave etc., 2014).
The content of the invention
In view of the problems of above-mentioned prior art, the present invention provides a kind of medical composition, the medical composition
Comprising mitochondria specialized cell (MitoCell), the wherein mitochondria specialized cell is that the pretreated fat of Chinese angelica root extract is dry
Cell, makes fat stem cell just tend to the direction of class neuronal cell when external.Mitochondria specialized cell is administered to body
The ratio that class neuronal cell is divided into after interior increases.It is class god that medical composition of the present invention can be effectively improved in differentiation in vivo
Ratio and the caused immune response of reduction through cell, and reach the purpose for the treatment of Parkinson's disease.
The present invention provides a kind of culture medium of mitochondria specialized cell, and the culture medium includes Chinese angelica root extract.
In one embodiment of the invention, wherein, Chinese angelica root extract include butylidene phthalide
(butylidenephthalide)。
The present invention provides a kind of preparation method of mitochondria specialized cell, before being carried out to stem cell with Chinese angelica root extract
Treatment.
The present invention also provides a kind of mitochondria specialized cell, and the mitochondria specialized cell is through doing that Chinese angelica root extract is processed
Cell.
In one embodiment of the invention, wherein, mitochondria specialized cell be stem cell.
In one embodiment of the invention, wherein, mitochondria specialized cell be fat stem cell.
In one embodiment of the invention, wherein, according to JC-1 fluorescent staining results, the line grain of mitochondria specialized cell
The red green fluorescence ratio of body film potential is 6.5~2.7.
The present invention additionally provides a kind of medical composition for increasing class neuron, the medical composition includes 50%~90%
Mitochondria specialized cell.
Present invention simultaneously provides a kind of method for treating nerve degenerative diseases, the method is comprised the following steps：By line grain
Body specialized cell is given into individual brain.
Brief description of the drawings
Figure 1A-Figure 1B shows survival rate of the fat stem cell under the Chinese angelica root extract of various concentrations.
Fig. 2 shows that Nurr1, BDNF of class neurons secrete of the fat stem cell under the Chinese angelica root extract of various concentrations increase
Height, represents fat stem cell and the destiny trend for being divided into nerve cell is presented；And SDF1 risings then represent stem cell homing
(homing) ability increases, and IL-8 gene expressions decline then represents inflammatory response reduction.
The mitochondrial blue green fluorescence ratio of Fig. 3 A display mitochondria specialized cells is reduced, therefore, mitochondria specialized cell
(MitoCell) mitochondrial membrane potential is differed with the mitochondrial membrane potential of normal cell, and Fig. 3 B are presented this mitochondria specialization
The cell still necessary characteristic (CD44/CD105) with stem cell.
Fig. 4 A- Fig. 4 B show balance beam test result.Result shows, is giving fat stem cell (the 3rd group) or mitochondria
Mouse after specialized cell (MitoCell) (the 4th group) substantially increases balanced capacity, and the 4th group of effect of the effect than the 3rd group is good.
Fig. 5 shows rotating wheel test result.Result shows, is giving fat stem cell (the 3rd group) or mitochondria specialization is thin
Mouse after born of the same parents (MitoCell) (the 4th group) can recover the balance and the coordination ability of mouse, the 4th group of effect of the effect than the 3rd group
It is really good.
Fig. 6 A- Fig. 6 C show eight passage Shuttle box results, as a result show, give fat stem cell (the 3rd group) or
Mouse after mitochondria specialized cell (MitoCell) (the 4th group) can clearly be seen that the recovery of its capacity, the 4th group of effect
Effect of the fruit than the 3rd group is good.
Fig. 7 shows brain sections H＆E (Hematoxylin and Eosin) coloration result, as a result shows that fat is dry thin
After born of the same parents and the injection of mitochondria specialized cell give, any toxicity is had no to nerve cell and the inflammatory reaction of brain will not be promoted.
Specific embodiment
The present invention is on a kind of fusion protein of innovation, its immunoglobulin for including a bioactive molecule and part
Molecule.Each side of the present invention is related to the fusion protein disclosed by fusion protein, the composition comprising it and manufacture and use
Method.The fusion protein of exposure is applied to extend serum half-life of the biomolecule in biology.
Detailed description presented below implements the present invention to assist those skilled in the art.Those skilled in the art
In the case of being appreciated that the adjustment or change of embodiment explained below, the spirit or scope for being included without departing from here, quilt
It is contained in the present invention.The term used in present invention explanation is only for narration specific embodiment rather than is intended as this hair
Bright limitation.Paragraph heading used below is the limitation of the content for organized purpose rather than as explanation.
Radix Angelicae Sinensis can be dried by programs such as freeze-drying, spray drying, evaporation or heat dryings.In the present invention,
Radix Angelicae Sinensis can be main root, lateral root or fiber.Radix Angelicae Sinensis can be extracted using solvent, to obtain Chinese angelica root extract.For example, can be used super
Supercritical fluid extraction, water extraction or organic solvent extracting process.Chinese angelica root extract of the invention preferably contains butylidene phthalide
(butylidenephthalide)。
" stem cell " of the present invention refer to cell in the state of undifferentiated or partial differentiation, with self-renewing
Characteristic, and can have the potentiality of development for being divided into various kinds of cell type, without the meaning of the specific hint on potentiality of development.
" stem cell " includes embryonic stem cell or adult stem cell.Adult stem cell is isolated from various adult tissues, including blood, bone
Marrow, brain, olfactory epithelium, skin, pancreas, skeletal muscle and cardiac muscle.Stem cell of the invention include but is not limited to fat stem cell,
NSC, stem cell of neural crest, interstital stem cell, candidate stem cell, pancreas stem cell, skin progenitor cell, embryonic stem cell,
Endothelial stem cell, liver stem cells, gut epithelial stem cells and germline stem cell, preferably fat stem cell.
The invention provides mitochondria specialized cell.Mitochondria specialized cell of the invention is obtained by following treatment
：By medium treatment at least 1 hour of the stem cell containing Chinese angelica root extract of the invention and/or butylidene phthalide, preferably
It is more than 2,3,4,5,6,7,8,9,10,15,20 or 24 hours, more preferably more than 1,2,3,4,5,6,7,8,9 or 10 days.
It should be noted that mitochondria specialized cell through Chinese angelica root extract after processing, the still feature with stem cell.This
Outward, mitochondria specialized cell can be divided into class neuronal cell in Mice Body.
When stem cell is processed through Chinese angelica root extract, the mitochondria activation of cell, but the stem cell still has stem cell
Characteristic.For example, before treatment, after, it is all through process stem cells in can detect CD44+/CD105+ surface markers.
The present invention provides a kind of preparation method of mitochondria specialized cell, is included in culture medium and cultivates stem cell, wherein,
The culture medium contains Chinese angelica root extract.
It is used to prepare the medical composition for the treatment of DD the present invention additionally provides a kind of mitochondria specialized cell
Purposes, it is characterised in that by injection system, makes mitochondria specialized cell enter individual brain.
The present invention additionally provides a kind of medical composition for treating nerve degenerative diseases.Medical composition bag of the invention
Specialized cell containing mitochondria, the wherein medical composition comprising the mitochondria specialized cell that active dose is for about 50%~90%,
Optimum activity dosage is 80%~90% mitochondria specialized cell.
Medical composition of the present invention can be effectively facilitated the quantity and quality of brain neuroblastoma unit cell, to increase the flat of individuality
Weighing apparatus and the coordination ability.It is of the present invention individual for the mankind or non-human animal (e.g., mouse, dog, cat, sheep, ox, horse, monkey
Deng), the preferably mankind.
Importantly, mitochondria specialized cell of the invention can not only significantly increase the number of dopamine neuronal cell
Amount, it is often more important that, can also reduce the immune response given produced by mitochondria specialized cell.Mitochondria specialized cell is not better than
Stem cell through processing.
Medical composition of the invention can be administered alone, or the treatment method with other ofneurodegenerative sacred diseases
Or medicine combination.
In sum, mitochondria specialized cell of the invention can increase dopamine neuronal cell and (be particularly at midbrain black
Matter area), the effect with old nerve degeneration relevant disease (such as Parkinson's disease and Alzheimer's disease etc.) for the treatment of.This
Outward, by mitochondria specialized cell of the invention give in vivo produced by immunological rejection ratio by fat stem cell give to
Immune response produced by vivo is low.
The extra specific embodiment of the present invention includes but is not limited to following.
The culture and pre-treatment of the mitochondria specialized cell (MitoCell) of embodiment 1.
Mitochondria specialized cell (MitoCell) be fat stem cell is incubated in fat stem cell culture solution prepare and
Obtain, wherein, fat stem cell culture solution is extracted comprising Keratinocyte-SFM (1X) Liquid (Gibco), Niu Chuiti
Thing (Gibco), EGF (Gibco), N-acetyl-L-cysteine (Sigma), L-AA 2- phosphate magnesium hydrates
(Sigma), hyclone (HyClone) (10%), and the extraction of the Radix Angelicae Sinensis containing 0,5,10,20,40,80,160 and 320 μ g/ml
Thing, wherein comprising described butylidene phthalide.Additionally, mitochondria specialized cell as described below is being processed through Chinese angelica root extract
Fat stem cell afterwards.
Reference picture 1, after 24 hours of incubation, when Chinese angelica root extract concentration is more than 160 μ g/mL, mitochondria specialized cell
Survival rate declines.After 48 hours of incubation, when Chinese angelica root extract concentration is more than 80 μ g/mL, the survival rate of mitochondria specialized cell
Can decline.
In addition, fat stem cell to be incubated at the Radix Angelicae Sinensis extraction of 0,0.3125,0.625,1.25,2.5,5 and 20 μ g/mL respectively
In taking thing.The expression quantity of Nurr1, BDNF, SDF1, IL-8 gene in mitochondria specialized cell is detected, and is used to find out most suitable
Close the dosage for the treatment of.
Reference picture 2, under the Chinese angelica root extract of high concentration (20 μ g/mL), the expression quantity of Nurr1, BDNF and SDF1 gene
Increase, but the expression of IL-8 genes can be suppressed.
Reference picture 3, the mitochondrial JC-1 dyeing red green Fluorescence Ratios of Fig. 3 A display mitochondria specialized cells change.It is special
The electrical potential energy for changing the mitochondrial membrane of cell substantially changes；Fig. 3 B are shown by flow cytomery mitochondria specialized cell.
There is mitochondria specialized cell CD44+/CD105+ to express, and display mitochondria specialized cell still has stem cell properties.20 μ g/mL's
Chinese angelica root extract is chosen to carry out subsequent experimental.
Embodiment 2. induces the foundation of op parkinson's disease mouse model
With eight week old, the C57BL/6 male mices that about 25 grams or so of body weight are purchased makees experimental subjects.Mouse point cage is four
Laundering period a couple of days is given after group, it is to avoid mouse is carried out and result because of urgent, anxiety influence experiment.The previous day tested, first
Carry out neuroethology observation analysis.Operation consent gives the chloraldurate of mouse 4% (cholra hydrate) for 10 minutes, and dosage is
1mL/g/kg body weight.Mouse Weight can give 0.25 milliliter of 4% chloraldurate every time about all at 25 grams.Additionally, giving
Isoflurane (isoflurane) is given with anesthetized mice, it is to avoid mouse revives during performing the operation.
By MPTP (MPTP) with physiological saline water dissolves in mouse
Parkinson's disease is induced, via conversion, every mouse gives MPTP in the way of intraperitoneal injection (I.P.), injects 4 times within one day,
Every 2 hours, dosage was 20mg/kg.And with 1x106The experiment group of individual cell/be often only given Parkinson's disease, as shown in table 1.
(1) the 1st group：Control group, does not inject MPTP；
(2) the 2nd groups：Negative control group, physiological saline is given after injection MPTP induction Parkinson's diseases to brain；
(3) the 3rd groups：Experimental group, 1x10 is given after injection MPTP induction Parkinson's diseases to brain6Individual fat stem cell；
(4) the 4th groups：Experimental group, 1x10 is given after injection MPTP induction Parkinson's diseases to brain6Individual mitochondria specialization is thin
Born of the same parents.
Embodiment 3. is performed the operation the analysis of front/rear neuroethology
3.1 balance beams are tested
Balance beam test (Beam walking analysis) is used for measuring the balanced capacity of mouse, by mouse put as
On the balance beam of 80cm, the record number of times that is slipped out by the time and the rear foot of balance beam of mouse, with analyze the balance of mouse with
Coordinate.Testing time is 60 seconds, if still cannot still be calculated with 60 seconds by balance beam more than 60 seconds mouse.
Reference picture 4A, after mouse injects MPTP to induce Parkinson's disease (the 2nd group), it is impossible to complete balance beam test.
Give after fat stem cell or mitochondria specialized cell (the 3rd group, the 4th group), hence it is evident that increase the balanced capacity of mouse.
Reference picture 4B, after mouse injects MPTP to induce Parkinson's disease (the 2nd group), the number of times that it is fallen increases.Giving
Give after fat stem cell or mitochondria specialized cell (the 3rd group, the 4th group), hence it is evident that reduce the number of times that mouse falls.
Found by experimental result, mitochondria specialized cell (the 3rd group) is compared to fat stem cell (the 4th group) to the mouse that gives
With preferably therapeutic effect.
3.2 rotating wheels are tested
Rotating wheel test (Rotarod analysis) is equally used for measuring the balance and the coordination ability of mouse, in test
The last week, mouse will be put to roller training by we, until time of the mouse on roller can perform the operation more than three minutes
Afterwards, the balanced capacity recovered after rotating wheel test assessment mouse treatment is recycled, the speed of roller is 5rpm.
Reference picture 5A, Fig. 5 B, after injection MPTP is to induce Parkinson's disease (the 2nd group), it balances and coordinates energy mouse
Power is decreased obviously.However, after fat stem cell or mitochondria specialized cell is given (the 3rd group, the 4th group), mouse can be recovered
Balance and the coordination ability, preferably give the mouse (the 4th group) of mitochondria specialized cell.
3.3 8 passage Shuttle box (Locomotor activity box)
Before being monitored, first put mouse into sampling box wait and adapt it to for 10 to 20 minutes.Eight passage shuttle box connect
Computer is connect, is shown and is recorded mouse via sensing and walked about (horizontal displacement), and came back or climb (vertical position during 30 minutes
Move) and overall total displacement data, the entire data that converges afterwards carries out statistical analysis.
Reference picture 6A, Fig. 6 B, Fig. 6 C, after injection MPTP is to induce Parkinson's disease (the 2nd group), it is balanced and association mouse
Tune ability is decreased obviously.However, after fat stem cell or mitochondria specialized cell is given (the 3rd group, the 4th group), can recover small
Mouse vertically moves number of times (Fig. 6 A), time (Fig. 6 B) and ability (Fig. 6 C), preferably gives the mouse of mitochondria specialized cell
(the 4th group).
Mouse is used the sacrifice method (2 to 3 times of anaesthesia dosages) of excessive anesthesia, waits mouse deep anaesthesia, starts profit
Perfusion is carried out with salt solution, it is after blood is all gone out then stiff to four limbs with metaformaldehyde (paraformaldehyde) perfusion, enter
The action that row brain takes out.
Skin behind ear is cut off with sharp scissors straight, then skin is vertically cut off overhead, to expose skull,
Cut off above mouse neck with scissors, neck is broken with cerebellum portion connection, point of a knife is carefully goed deep into skull to nose
Cut off, be careful not to hurt brain.Emptied under brain, take out whole brain, and by the dehydration of mouse brain, be placed on cut brain afterwards
On platform, cut before this brain platform can be firstly placed on it is pre- cool to avoid brain injury on ice.
Cerebellum and olfactory bulb in removal brain, two parts are cut into by the brain left and right sides, carry out OCT (optimized cutting temperature；
Optimal cutting temperature) embedding, then obtain mouse brain section using freezing-microtome.
Mouse brain section H＆E will be obtained to dye, the brain cell of observation injection fat stem cell and mitochondria specialized cell
Whether there is impaired or inflammatory reaction, as a result show that brain cell occurs (Fig. 7) without inflammatory response is undermined.
Claims (15)
1. it is a kind of cultivate stem cell culture medium, wherein, the culture medium include Chinese angelica root extract.
2. culture medium as claimed in claim 1, wherein, the Chinese angelica root extract is butylidene phthalide.
3. culture medium as claimed in claim 2, wherein, the concentration of the Chinese angelica root extract is 5~160 μ g/ μ l.
4. culture medium as claimed in claim 2, wherein, the concentration of the Chinese angelica root extract is 20 μ g/ μ l.
5. a kind of preparation method of mitochondria specialized cell, methods described is included in the culture medium described in claim 1 and cultivates
Cell.
6. preparation method as claimed in claim 1, wherein, the stem cell is selected from by following constituted group：Fat is dry
Cell, NSC, stem cell of neural crest, interstital stem cell, candidate stem cell, pancreas stem cell, skin progenitor cell, embryo do
Cell, endothelial stem cell, liver stem cells, gut epithelial stem cells and germline stem cell.
7. a kind of mitochondria specialized cell, wherein, the mitochondria specialized cell is derived from the stem cell processed through Chinese angelica root extract；
Wherein, the mitochondrial JC-1 dyeing red green fluorescence ratios of the mitochondria specialized cell are 6.5~2.7.
8. mitochondria specialized cell as claimed in claim 7, wherein, the stem cell is selected from by following constituted group：
Fat stem cell, NSC, stem cell of neural crest, interstital stem cell, candidate stem cell, pancreas stem cell, skin progenitor cell,
Embryonic stem cell, endothelial stem cell, liver stem cells, gut epithelial stem cells and germline stem cell.
9. mitochondria specialized cell as claimed in claim 7, wherein, the mitochondria activity of the mitochondria specialized cell changes
Become, and the mitochondria specialized cell has the cell marking Nuur1 that altimeter reaches.
10. mitochondria specialized cell as claimed in claim 7, wherein, the mitochondria specialized cell has low expression
Cell marking IL8.
11. mitochondria specialized cells as claimed in claim 7, wherein, the mitochondria specialized cell expresses stem cell labeling
CD44+ and CD105+.
A kind of 12. medical compositions for treating nerve degenerative diseases, the medical composition is comprising as claimed in claim 7
Mitochondria specialized cell and pharmaceutically acceptable salt.
13. medical compositions as claimed in claim 11, wherein, the medical composition includes 50%~90% mitochondria
Specialized cell.
14. medical compositions as claimed in claim 11, wherein, the medical composition includes 80%~90% mitochondria
Specialized cell.
A kind of 15. mitochondria specialized cells are used for the purposes of the medical composition for preparing treatment nerve degenerative diseases, wherein, institute
It is the stem cell processed through Chinese angelica root extract to state mitochondria specialized cell.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202011304171.5A CN112646774B (en) | 2014-09-11 | 2015-09-11 | Pharmaceutical composition for treating neurodegenerative diseases with mitochondria-specific cells |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| US201462049090P | 2014-09-11 | 2014-09-11 | |
| US62/049,090 | 2014-09-11 | ||
| PCT/CN2015/089402 WO2016037587A1 (en) | 2014-09-11 | 2015-09-11 | Pharmaceutical compositions for treating degenerative neurological disease with mitocells |
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| CN202011304171.5A Division CN112646774B (en) | 2014-09-11 | 2015-09-11 | Pharmaceutical composition for treating neurodegenerative diseases with mitochondria-specific cells |
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| CN106795490B CN106795490B (en) | 2021-01-19 |
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| CN202011304171.5A Active CN112646774B (en) | 2014-09-11 | 2015-09-11 | Pharmaceutical composition for treating neurodegenerative diseases with mitochondria-specific cells |
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| US (1) | US20160074438A1 (en) |
| CN (2) | CN106795490B (en) |
| TW (1) | TWI707040B (en) |
| WO (1) | WO2016037587A1 (en) |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| TWI675678B (en) * | 2018-08-23 | 2019-11-01 | 國為生醫科技股份有限公司 | Use of n-butylidenephthalide in dopaminergic progenitor cell transplantation |
| CN111494412A (en) * | 2019-01-30 | 2020-08-07 | 台湾粒线体应用技术股份有限公司 | Use of isolated exogenous mitochondria for the preparation of a composition for the treatment of multiple system degeneration and its symptoms |
| CN111759861A (en) * | 2019-03-27 | 2020-10-13 | 台湾粒线体应用技术股份有限公司 | Use of mitochondria for the preparation of a pharmaceutical composition for the treatment and/or prevention of alzheimer's disease |
| CN112601532A (en) * | 2018-07-22 | 2021-04-02 | 米诺维亚疗法有限公司 | Mitochondrion-enhanced therapy using stem cells enriched for functional mitochondria |
| CN113801836A (en) * | 2020-06-16 | 2021-12-17 | 台湾粒线体应用技术股份有限公司 | Culture method for improving mitochondrial function and composition used by same |
Families Citing this family (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106190963A (en) * | 2016-07-13 | 2016-12-07 | 浙江大学 | A kind of method using mitochondrial transplantation to promote injured neuron survival |
| TWI608839B (en) * | 2016-11-04 | 2017-12-21 | 國璽幹細胞應用技術股份有限公司 | Composition for treating tendonitis and manufacture thereof |
| TWI781322B (en) * | 2019-05-27 | 2022-10-21 | 台灣粒線體應用技術股份有限公司 | A culture composition and an use for improving the function of mitochondria thereof |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2010110755A1 (en) * | 2009-03-27 | 2010-09-30 | Moleac Pte. Ltd. | Therapy for promoting cell growth |
| CN103864947A (en) * | 2014-03-17 | 2014-06-18 | 重庆医科大学 | Process for separating and extracting angelica polysaccharide and application thereof |
Family Cites Families (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2003078967A2 (en) * | 2002-03-12 | 2003-09-25 | Oregon Health & Science University | Stem cell selection and differentiation |
| CN1424396A (en) * | 2002-12-13 | 2003-06-18 | 中山大学 | Preparation of implanting neurons for treating nerve injury diseases |
| WO2005040362A1 (en) * | 2003-10-29 | 2005-05-06 | Fcb Pharmicell Co., Ltd | Method for differentiating mesenchymal stem cell into neural cell and pharmaceutical composition containing the neural cell for neurodegenerative disease |
| TW200609355A (en) * | 2004-09-01 | 2006-03-16 | Tzu Chi Buddhist General Hospital | Nurr1-positive neuron stem cells, pharmaceutical composition thereof and methods for their isolation, culture and preservation |
| KR20080015802A (en) * | 2005-05-24 | 2008-02-20 | 디에스엠 아이피 어셋츠 비.브이. | Ligustilide derivatives for the treatment of inflammatory disorders |
| KR100679642B1 (en) * | 2005-11-16 | 2007-02-06 | 주식회사 알앤엘바이오 | Multipotent stem cell derived from human adipose tissue and cellular therapeutic agents comprising the same |
| SG177238A1 (en) * | 2010-07-22 | 2012-03-29 | Affiliated Hospital Of Ningxia Medical University | A method of producing neurons from stem cells, the neurons and uses thereof |
| TWI484033B (en) * | 2013-01-25 | 2015-05-11 | Univ China Medical | Method and kit for culturing stem cells |
| US20140271568A1 (en) * | 2013-03-12 | 2014-09-18 | Hawking Biological Technology Co., Ltd | Method and kit for providing an increased expression of telomerase, brain-derived neurotrophic factor, stromal cell-derived factor-1, cxc chemokine receptor 4, and/or immune regulatory factor of stem cell |
| US9526679B2 (en) * | 2014-08-28 | 2016-12-27 | Aphrozone Co., Ltd. | Method and apparatus of manufacturing cosmetic products for regenerating skin cell |
-
2015
- 2015-09-11 US US14/851,402 patent/US20160074438A1/en not_active Abandoned
- 2015-09-11 CN CN201580043621.0A patent/CN106795490B/en active Active
- 2015-09-11 CN CN202011304171.5A patent/CN112646774B/en active Active
- 2015-09-11 WO PCT/CN2015/089402 patent/WO2016037587A1/en active Application Filing
- 2015-09-11 TW TW104130090A patent/TWI707040B/en active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2010110755A1 (en) * | 2009-03-27 | 2010-09-30 | Moleac Pte. Ltd. | Therapy for promoting cell growth |
| EP2411028A1 (en) * | 2009-03-27 | 2012-02-01 | Moleac Pte Ltd. | Therapy for promoting cell growth |
| CN103864947A (en) * | 2014-03-17 | 2014-06-18 | 重庆医科大学 | Process for separating and extracting angelica polysaccharide and application thereof |
Non-Patent Citations (3)
| Title |
|---|
| QIAOZHI WANG等: "Differentiation of human adipose-derived stem cells into neuron-like cells by Radix Angelicae Sinensis", 《NEURAL REGEN RES》 * |
| SHIH-PING LIU等: "n-Butylidenephthalide (BP) Maintains Stem Cell Pluripotency by Activating Jak2/Stat3 Pathway and Increases the Efficiency of iPS Cells Generation", 《PLOS ONE》 * |
| 王巧稚等: "当归和丁酸羟基茴香醚诱导人脂肪干细胞向神经元样细胞分化的研究", 《药学分析杂志》 * |
Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112601532A (en) * | 2018-07-22 | 2021-04-02 | 米诺维亚疗法有限公司 | Mitochondrion-enhanced therapy using stem cells enriched for functional mitochondria |
| TWI675678B (en) * | 2018-08-23 | 2019-11-01 | 國為生醫科技股份有限公司 | Use of n-butylidenephthalide in dopaminergic progenitor cell transplantation |
| CN111494412A (en) * | 2019-01-30 | 2020-08-07 | 台湾粒线体应用技术股份有限公司 | Use of isolated exogenous mitochondria for the preparation of a composition for the treatment of multiple system degeneration and its symptoms |
| CN111759861A (en) * | 2019-03-27 | 2020-10-13 | 台湾粒线体应用技术股份有限公司 | Use of mitochondria for the preparation of a pharmaceutical composition for the treatment and/or prevention of alzheimer's disease |
| CN111759861B (en) * | 2019-03-27 | 2023-01-13 | 台湾粒线体应用技术股份有限公司 | Use of mitochondria for the preparation of a pharmaceutical composition for the treatment and/or prevention of alzheimer's disease |
| CN113801836A (en) * | 2020-06-16 | 2021-12-17 | 台湾粒线体应用技术股份有限公司 | Culture method for improving mitochondrial function and composition used by same |
Also Published As
| Publication number | Publication date |
|---|---|
| TWI707040B (en) | 2020-10-11 |
| TW201610157A (en) | 2016-03-16 |
| CN112646774A (en) | 2021-04-13 |
| CN112646774B (en) | 2024-05-17 |
| WO2016037587A1 (en) | 2016-03-17 |
| US20160074438A1 (en) | 2016-03-17 |
| CN106795490B (en) | 2021-01-19 |
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